Treatment for hypercholesterolemia

ABSTRACT

The present invention relates to a method of modifying serum cholesterol levels in a mammal by topically administering to the skin of the mammal an effective amount of at least one phospholipid. Atherosclerosis, and related complications, can be treated, or prevented, using the present method.

BACKGROUND OF THE INVENTION

1. Technical Field

The present invention relates, in general, to hypercholesterolemia and,more specifically, to the use of phospholipids in a method of modifyingblood cholesterol levels.

2. Background Information

Elevated blood cholesterol level is a major risk factor for thedevelopment of coronary heart disease, stroke and peripheral vasculardisease. Of these, coronary heart disease has remained the leading causeof death in the United States and other affluent countries, in spite ofrecent advances in the management of the disease and understanding ofthe disease mechanism. It is understood that the underlying cause of theabove-mentioned diseases is atherosclerosis, an insidious process ofdeposition of cholesterol and its esters, compounded with othermaterials, in characteristic plaques in the arterial wall.

Lowering blood cholesterol can reduce the risk of coronary heart disease(Stamler et al., JAMA 256:2823, 1986; The Lipid Research ClinicsCoronary Primary Prevention Trial Results, I: Reduction in incidence ofcoronary heart disease. JAMA 251:351, 1984; The Lipid Research ClinicsCoronary Primary Prevention Trial Results, II: The relation of reductionin incidence of coronary heart disease to cholesterol lowering. JAMA251:365, 1984). In fact, each one percent reduction in blood cholesterollevel is believed to result in approximately a two percent reduction incoronary heart disease rate (National Institutes of Health, ConsensusDevelopment Conference Statement. Vol. 5, No. 7).

Recommended methods for lowering blood cholesterol include reducingweight, increasing exercise, and altering diet by lowering cholesterolintake and substituting saturated fat with polyunsaturated fat. Theeffectiveness and drawbacks of several drugs that have been approved byFDA for use in the treatment of hypercholesterolemia have been recentlyreviewed (Blum et al., JAMA 261:3582, 1989).

Cholesterol and phospholipids are essential components of cellularorganelles and membranes of animals. They are also major components oflipoproteins of the circulating blood. Their simultaneous occurrence inanimal tissues indicates an affinity between molecules of phospholipidsand cholesterol. This affinity has been utilized in the technology forpreparing liposomes. Liposomes prepared from phospholipids, which arenotoriously unstable, can be stabilized by the addition of cholesterol.

The affinity of phospholipids for cholesterol provides a basis for thehypothesis that phospholipids, when properly administered, could removecholesterol from atherosclerotic plaques, and thus reduce the risk forcoronary heart disease. Indeed, in experimental animals, intravenousadministration of phospholipids has resulted in resolution ofatherosclerotic lesions (Friedman et al., Proc. Soc. Exp. Biol. Med.95:580, 1957; Howard et al., Atherosclerosis 14:17, 1971; Stafford etal., Artery 1(2):105, 1975). Extensive changes in serum lipoproteins inrabbits after intravenous injection of liposomes made of egg yolkphospholipids have been demonstrated. Such changes may haveanti-atherogenic effects (Mendez et al., Lipids 23:961, 1988).

Although the above-described results suggest potential usefulness ofphospholipids in the prevention and treatment of atherosclerosis,intravenous injection is an invasive method of drug delivery, and hasobjectionable features The development of a more convenient method fordelivering phospholipids, and thereby effecting a reduction in bloodcholesterol levels, is needed. The present invention provides suchmethod.

SUMMARY OF THE INVENTION

It is a general object of the invention to provide a convenient methodfor effecting modification of serum cholesterol levels.

It is a specific object of the invention to provide a method ofpreventing and treating atherosclerosis, and related complications.

Various other objects and advantages of the present invention willbecome obvious from the drawings and description of the invention thatfollow.

The present invention relates to a method of lowering serum cholesterollevels in a mammal. The method comprises topically administering to amammal an amount of a phospholipid-containing composition sufficient toreduce serum cholesterol to an acceptable level.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the response in serum cholesterol level to the topicalapplication of a phospholipid solution to the skin of 5 rabbits.

FIG. 2 shows the response in LDL cholesterol level to the topicalapplication of a phospholipid solution to the skin of 5 rabbits.

FIG. 3 shows the response in HDL cholesterol level to the topicalapplication of a phospholipid solution to the skin of 5 rabbits.

FIG. 4 summarizes the effects of topical application of a phospholipidsolution on cholesterol levels.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to a method of lowering serum cholesterollevels in mammals. The method comprises topically administering to themammal an amount of a phospholipid-containing composition sufficient tolower serum cholesterol to an acceptable level. The present invention isfurther directed to a method of preventing and treating atherosclerosis,and related complications, by topically administering to a mammal aphospholipid-containing composition in an amount sufficient to lower theserum cholesterol level of the mammal. The methods of the presentinvention result in the lowering of serum LDL cholesterol levels and theenhancing of serum HDL cholesterol levels in the mammal undergoingtreatment.

Phospholipid-containing compositions suitable for use in the presentmethod comprise at least one phospholipid in a concentration of between,for example, 5% and 50% (w/v). Any natural or synthetic phospholipid,lecithin (phosphatidylcholine), phosphatidylethanolamine andphosphatidylserine, having an affinity for cholesterol can be used inthe composition, however, lecithin is preferred.

Also included in compositions suitable for use in the present inventionis a pharmaceutically acceptable carrier The carrier is, advantageously,an alcohol, such as ethanol, however, any carrier that does not affectthe affinity of the phospholipid for cholesterol can be used. Otherpossible carriers include isopentenyl, isobutyl or isopropyl alcohol.

One of ordinary skill in the art can readily determine the appropriateamount of the phospholipid-containing composition to be administered toa particular patient to achieve the appropriate reduction in serumcholesterol level. In one embodiment, about 25 ml of a 25%phospholipid/ethanol solution is applied 1 or 2 times per week.

The phospholipid-containing composition to be used in the present methodcan be rubbed onto or sprayed onto the skin of the individual whoseserum cholesterol is sought to be altered. Alternative forms of topicaladministration can also be used. For example, thephospholipid-containing composition can be present in a patch which canbe adhered to the skin of the individual in a manner such that topicaladministration is effected.

In addition to active ingredient (phospholipid) and carrier, otheringredients that confer desirable characteristics on the composition canalso be included in the formulation (i.e., perfumes, coloring agents,water, absorption enhancers, etc.).

The present invention is described in further detail in the followingnon-limiting Examples.

EXAMPLE I

New Zealand white rabbits usually have a serum cholesterol level in therange of 30-50 mg/dl. By chance, an experimental rabbit (R4)spontaneously developed hypercholesterolemia, with a serum cholesterollevel of 110 mg/dl.

New Zealand white rabbits were fed normal rabbit chow and maintained ata controlled temperature with a 12 hour dark/light cycle. Food wasremoved from the rabbits at least 12 hours prior to collection of bloodsamples from the ear vein.

Phospholipid was extracted from egg yolk by standard procedures andpurified. The phospholipid solution was prepared by adding ethanol tothe purified phospholipids to a final concentration of 25-50% (w/v).

To test whether topically applied phospholipids can lower serumcholesterol, the backs of R4 and four other rabbits (R1, R2, R3 and R5)were shaved, and 3 ml of a solution of 25% (w/v) egg yolk phospholipidsin ethanol were applied to the shaved skin of R3, R4 and R5, while 3 mlof ethanol alone was applied to the controls (R1 and R2). Blood sampleswere taken weekly from all rabbits, 2 weeks before the application and 6weeks thereafter, and again at the end of the 10th week. Cholesterol wasassayed in serum, high density lipoproteins (HDL) and low densitylipoproteins (LDL) by established procedures. The results are shown inFIGS. 1-3.

The data show that in the control animals (R1 and R2) cholesterol levelsheld steady throughout the experimental period. In contrast, data fromR4 show dramatic drops in serum and LDL cholesterol, and also anincrease of HDL, 3 weeks after topical application of phospholipids.

The response from R5 whose serum cholesterol was slightly elevated, wasless dramatic. The data show little or no effect on serum cholesteroland lipoproteins in R3 which had low serum cholesterol.

The above experiment was repeated with 7 more rabbits having serumcholesterol levels above 100 mg/dl. These animals were selected from 120rabbits after their serum cholesterol levels were measured.

The results of the 7 rabbits after topical treatment with phospholipidsare shown in FIG. 4. A decrease in serum and LDL cholesterol and anincrease in HDL cholesterol were observed 3 weeks after the treatment.

The decrease in serum cholesterol, and especially the decrease in LDLcholesterol, are thought to be desirable in terms of the prevention ofatherosclerosis. The increase of HDL cholesterol, which has been labeledas "good cholesterol" in the lay press, is of importance not only inreducing the risk of atherosclerosis but also for reversal of thedisease process. These findings form the basis for the prevention andtreatment of atherosclerotic diseases in humans by topical treatmentwith phospholipids.

EXAMPLE II

RJH, a 62 year old man who was in apparent good health, although havinga long history of elevated serum cholesterol and triglyceride levels,volunteered to test the effect of topical lecithin on serum cholesterol.A fasting blood sample was drawn on day 1 and on day 8 for themeasurements of serum cholesterol and triglycerides, and cholesterol inthe low density lipoproteins (LDL), very low density lipoproteins (VLDL)and high density lipoproteins (HDL) by standard procedures. The resultsshowed that his serum lipid levels were steadily elevated (see table).In the evening of day 8, he applied 25 ml of a 50% solution of egglecithin (purified according to procedures published in the Journal ofthe American Oil Chemists Society (42:53-56 (1965)) in ethanol to covermost of his body surface. He experienced no ill effects in follow up.

Fasting blood samples were drawn from RJH on day 15 and day 23 for lipidmeasurements. The results showed decreases in serum and LDL cholesterolafter the treatment with topical lecithin. The 16.8% drop of LDLcholesterol from 191 to 159 mg/dl in 2 weeks after the treatment wasespecially remarkable. The data are summarized in the following table:

    ______________________________________                                        RJH LIPID PROFILE                                                             (mg/dl of serum)                                                              DAY      SC      TG        HDL   VLDL    LDL                                  ______________________________________                                         1       250     241       49    11      190                                   8       254     156       49    14      191                                   8       Treatment with topically applied lecithin.                           15       246     172       52    10      184                                  23       234     188       51    24      159                                  ______________________________________                                    

Abbreviations are: SC, Serum cholesterol; TG, serum triglycerides; HDL,high density lipoproteins; VLDL, very low density lipoproteins; LDL, lowdensity lipoproteins.

EXAMPLE III

BK, a 39 year old man who was in apparent good health although slightlyoverweight, became concerned about his elevated serum cholesterol level.Fasting blood samples were taken in three consecutive weeks, and theresults showed serum cholesterol consistently above 290 mg/dl. He wasgiven the following instruction:

1) Take a hot shower in the evening, and dry your skin with a towel.

2) Apply the solution given to you to your skin and rub it in well, tocover your legs, arms, the abdomen and the back. Use all the solution,and spread it to cover all your body.

3) To enhance percutaneous absorption, wrap your legs with a thinplastic film, e.g. Saran Wrap, and fix the wrapping with a tape or anAce bandage.

4) Rest or take a nap, and wait for at least 2 hours before washing offthe unabsorbed material on your skin.

The solution given him for each application was 12.5 ml of FormulaM-23-90 which has the following composition:

    ______________________________________                                        Formula M-23-9 (Total volume 78 ml)                                           ______________________________________                                        Soy lecithin   25 gm                                                          BHT            0.2 gm                                                         Ethanol        25 ml                                                          Isopropanol    18 ml                                                          Distilled water                                                                              18 ml                                                          ______________________________________                                    

BK returned each week or 10 days for analysis of fasting blood lipidprofile. His serum cholesterol and low density lipoproteins showeddefinite reductions after each treatment with Formula M-23-90. Hereported no undesirable effects. A summary of his lipid profiles isshown in the following table

    ______________________________________                                        BK LIPID PROFILE                                                              (mg/dl of serum)                                                              Day  SC         TG       HDL       VLDL  LDL                                  ______________________________________                                         1   292        356      34        98    160                                   9   295        311      42        108   145                                  16   294        167      40        56    198                                  16   Treatment with Formula M-23-90                                           24   262        263      35        52    175                                  24   Treatment with Formula M-23-90                                           35   242        248      37        81    124                                  35   Treatment with Formula M-23-90                                           42   236        257      38        72    126                                  ______________________________________                                    

Abbreviations are: SC, serum cholesterol; TG, serum triglycerides; HDL,high density lipoproteins; VLDL, very low density lipoproteins; LDL, lowdensity lipoproteins.

The entire contents of all references cited hereinabove are herebyincorporated by reference.

While the foregoing invention has been described in some detail forpurposes of clarity and understanding, it will be clear to one skilledin the art from a reading of this disclosure that various changes inform and detail can be made without departing from the true scope of theinvention.

What is claimed is:
 1. A method of increasing the ratio of high densitylipoprotein to low density lipoprotein in the serum of a mammal, whichcomprises topically administering to said mammal an amount of aphospholipid sufficient to increase said ratio.
 2. The method accordingto claim 1, wherein said ratio is increased by lowering the level of lowdensity lipoprotein cholesterol.
 3. The method according to claim 1,wherein said ratio is increased by enhancing the level of high densitylipoprotein cholesterol.
 4. The method according to claim 1, whereinsaid phospholipid is lecithin.
 5. The method according to claim 1,wherein said phospholipid is phosphatidylserine.
 6. The method accordingto claim 1, wherein said phospholipid is phosphatidylethanolamine. 7.The method according to claim 1, wherein said phospholipid is present asa liquid solution.
 8. The method according to claim 7, wherein saidphospholipid is present as an alcohol solution.
 9. The method accordingto claim 8, wherein said alcohol is ethanol.
 10. The method according toclaim 1, wherein said phospholipid is present in a composition havingthe form of a lotion, cream, gel or ointment.
 11. The method accordingto claim 1 wherein said phospholipid is soy lecithin administered incombination with butylated hydroxytoluene, ethanol, isopropanol anddistilled water.
 12. A transdermal patch comprising a reservoircontaining an amount of a phospholipid sufficient to increase the ratioof high density lipoprotein to low density lipoprotein in the serum of amammalwherein said patch is constructed such that said phospholipid isavailable for transdermal administration to a mammal wearing said patch.13. A method of lowering the serum cholesterol level of a mammal whichcomprises typically administering to said mammal an amount of aphospholipid sufficient to effect said lowering.
 14. A method oftreating atherosclerosis, and related complications, which comprisestopically administering to a patient in need of such treatment an amountof a phospholipid sufficient to treat atherosclerosis and said relatedcomplications.